Mixed Stimuli-Responsive Magnetic and Gold Nanoparticle System for Rapid Purification, Enrichment, and Detection of Biomarkers

A new diagnostic system for the enrichment and detection of protein biomarkers from human plasma is presented. Gold nanoparticles (AuNPs) were surface-modified with a diblock copolymer synthesized using reversible addition-fragmentation chain transfer (RAFT) polymerization. The diblock copolymer contained a thermally responsive poly(N-isopropylacrylamide) (pNIPAAm) block, a cationic amine-containing block, and;1 semi telechelic PEG(2)-biotin end group. When a mixed suspension of 23 nm pNIPAAm-modified AuNPs was heated with pNIPAAm-coated 10 nm iron oxide magnetic nanoparticles (mNPs) in human plasma, the thermally responsive pNIPAAm directed the formation of mixed AuNP/mNP aggregates that could be separated efficiently with magnet. Model studies showed that this mixed nanoparticle system could efficiently purify and strongly enrich the model biomarker protein streptavidin in spiked human plasma. A 10 ng/mL, streptavidin sample was mixed with the biotinylated pNIPAAm-modified AuNPs and magnetically separated in the mixed nanoparticle system with pNIPAAm mNPs. The aggregates were concentrated into a 50-fold smaller fluid volume at room temperature where the gold nanoparticle reagent redissolved with the streptavidin target still bound. The concentrated gold-labeled streptavidin could be subsequently analyzed directly using lateral flow immunochromatography. This rapid capture and enrichment module thus utilizes the mixed stimuli-responsive nanoparticle system to achieve concentration of a gold-labeled biomarker that can be directly analyzed using lateral flow or other rapid diagnostic strategies.